The working paradigm of the section is that there is a genetic basis to human disease and that understanding the genetic basis of disease will foster development of better diagnostic and treatment strategies. To this end we are studying a number of Mendelian diseases to identify the underlying gene defect and to understand how the product(s)of this/these gene mutation(s)result in abnormal development or disease. Specific disease conditions studied include: Dentinogenesis Imperfecta (DI): Dentin, the most abundant tissue in teeth is produced by odontoblasts which differentiate from mesenchymal cells in the dental papilla. Genetic mutations of the DSSP gene are responsible for dentinogenesis imperfecta and dentin dysplasia. We have identified mutations in the DSSP in individuals affected with DI and DD. Certain DSSP mutations are the result of defective transport of DSSP gene products. These findings have provided a basis to evaluate strategies to remove defective protein from the odontoblasts, and evaluation of intervention strategies. [unreadable] Amelogenesis Imperfecta (AI): We continue to study families segregating the amelogenesis imperfecta phenotype. Autosomal dominant and recessive families are being evaluated to identify the chromosomal locations of genetic mutations responsible for various forms of AI. Studies of dominant forms of the disease indicate a gene of major effect for AI on chromosome 8 in some families, while other families show no evidence for linkage to known AI genes, indicating additional genetic heterogeneity for the condition. [unreadable] Tricho-dento-osseous syndrome (TDO): Previously we determined that a DLX3 truncation mutation is responsible for TDO in humans. One feature of the condition is increased bone thickness and density. Our goal is to understand how this mutation results in these favorable osseous findings. Results of in vitro studies indicate that mutant Dlx3 may accelerate differentiation of osteoprogenitor cells. To evaluate this observation, we generated transgenic (TG) mice that carry the same deletion mutation. Transgenic mice have phenotypic features including taurodontism as well as enhanced bone density and thickness. These TG mice show defects in odontoblasts and cementoblasts resulting in reduced dentin formation. Bone density and thickness are markedly enhanced in TG mice. [unreadable] Hereditary gingival fibromatosis (HGF): We have previously identified a mutation of the son of sevenless 1 (SOS1) gene HGF. Fibroblasts with the mutation have higher proliferation rates, resulting in increased cell numbers and collagen. Using ectopic expression of wild-type and mutant SOS1 constructs, we found that truncated SOS1 could localize to the plasma membrane, without growth factor stimuli, leading to sustained activation of Ras/MAPK signaling. Additionally, we observed an increase in the magnitude and duration of ERK signaling in HGF gingival fibroblasts that was associated with phosphorylation of retinoblastoma tumor suppressor protein and the up-regulation of cell cycle regulators, including cyclins C, D, and E and the E2F/DP transcription factors. These factors promote cell cycle progression, and their up-regulation may underlie the increased gingival fibroblast proliferation observed. These findings elucidate the mechanisms for gingival overgrowth mediated by SOS1 gene mutation in humans. [unreadable] Papillon Lefvre syndrome (PLS): Mutations of the cathepsin C gene (CTSC)and resultant inactivity of cathepsin C enzyme are responsible PLS and several allelic conditions. Inactivity of cathepsin C has been demonstrated to result in the inability to activate neutrophil serine proteases (NSP). We have previously related inactivity of CTSC and NSP with dysregulation of MIP-1alpha, and suggested this may be important in the inability to regulate leukocyte at sites of inflammation in PLS, possibly contributing to tissue destruction. Because elevated levels of the macrophage inflammatory protein-1alpha (MIP-1alpha) are reported in inflammatory bone diseases including periodontitis, we evaluated the ability of interleukin-1beta (IL-1beta) and bacterial lipopolysaccharides (LPSs) to modulate MIP-1alpha expression in epithelial cells, fibroblasts, and polymorphonuclear leukocytes (PMNs). We found that MIP-1alpha expression in PMNs and gingival epithelial cells was induced by IL-1beta and LPS, but neither induced MIP-1alpha expression in gingival fibroblasts or osteoblastic cells. MIP-1alpha was highly expressed in the basal epithelial layer of inflamed gingiva but not in healthy gingiva. We also found that MIP-1alpha induced osteoclast formation. These findings suggest that MIP-1alpha expression by gingival epithelial cells may be important in initiating inflammation by facilitating accumulation and activation of leukocytes. The ability of MIP-1alpha to facilitate formation of multinuclear bone cells indicates a possible role in periodontitis-associated bone destruction. These findings indicate MIP-1alpha may play an important role in early and later stages of inflammatory-related periodontitis. We believe these findings have clinical implications for Mendelian and complex forms of periodontitis. [unreadable] To study gene-environment interactions that impact on oral diseases we have been evaluating the utility of incorporating microbial expression profiles and proteomic data into models that can be use to predict disease or to distinguish between diseased individuals and controls. We have assessed changes in the oral flora in intubated critical care patients, in an effort to identify changes in the oral flora that are associated with ventilation in hospital intensive care patients. Findings indicate observable changes in the oral flora, and characterization of the salivary proteome is currently underway. Studies of the salivary proteome led us to investigate changes in salivary proteome following allogeneic hematopoietic stem cell transplantation. As saliva contains many components of adaptive and innate immune response crucial for local host defenses, changes in salivary constituents could reflect systemic processes such as immune reconstitution and development of graft versus host disease(GVHD) that occur posttransplant. Serially collected saliva samples from patients undergoing allo-HCT were evaluated. Changes in salivary proteome were initially examined by SELDI-TOF mass spectrometry. Individual protein changes were identified by 2-dimensional differential in-gel electrophoresis (2D-DIGE) with subsequent MS/MS sequencing and ELISA. Significant increases and decreases in multiple salivary proteins that lasted at least 2 months posttransplant were detected by SELDI-TOF mass spectrometry. Lactoferrin and secretory leukocyte protease inhibitor demonstrated elevations 1 month post-HCT that persisted at least 6 months. Secretory IgA (sIgA) levels were decreased 1 month posttransplant, with recovery at approximately 6 months. Levels of salivary beta(2)-microglobulin were elevated at 6 months and correlated with sIgA levels. We conclude that Allo-HCT is associated with long-term changes in several salivary proteins important for innate immune responses. These results support further studies on the association of salivary proteins with posttransplant complications including infections and GVHD. We have also profiled the salivary proteome from individuals with infectious dental diseases, including caries, to determine if profiling the salivary proteome can add discriminatory power to models to identify individuals with or at risk for developing caries. This work is building on our previous observation that quantitative changes in groups of oral microbes can be related to the presence or absence of caries in groups of individuals.